The purpose of this study is to provide a culture for mouse bone marrow-derived macrophages (BMDM) and peritoneal macrophages (PM) and to characterize their molecular and cellular biology. The cell number and purity from the primary culture were assessed by cell counter and flow cytometry, respectively. Morphological features were evaluated by inverted microscope. Phagocytosis by macrophages was detected by the neutral red dye uptake assay. Phenotypic markers were analyzed by real-time fluorescent quantitative PCR. Our results show that the cell number was much higher from culture of BMDM than PM, while there was no significant difference regarding the percentage of F4/80+CD11b+ cells (98.30%±0.53% vs. 94.83%±1.42%; P>0.05). The proliferation rate of BMDM was significantly higher than PM in the presence of L929 cell conditioned medium, by using CCK-8 assay. However, PM appeared to adhere to the flask wall and extend earlier than BMDM. The phagocytosis capability of un-stimulated BMDM was significantly higher than PM, as well as lipopolysaccharide (LPS)-stimulated BMDM, except the BMDM stimulated by low dose LPS (0.1 μg/mL). Furthermore, Tnfα expression was significantly higher in un-stimulated BMDM than PM, while Arg1 and Ym1 mRNA expression were significantly lower than PM. The expression difference was persistent if stimulated by LPS+IFN-γ or IL-4. Our data indicate that bone marrow can get larger amounts of macrophages than peritoneal cavity. However, it should be aware that the molecular and cellular characteristics were different between these two culture systems.
Animals ; Bone Marrow Cells ; physiology ; Cells, Cultured ; Culture Media, Conditioned ; Lipopolysaccharides ; metabolism ; Macrophages ; classification ; physiology ; Mice ; Phagocytosis

This study was conducted to evaluate the impact of implementation of an automated liquid culture system on the diagnosis of tuberculous pleurisy in an HIV-uninfected patient population. We retrospectively compared the culture yield, time to positivity, and contamination rate of pleural effusion samples in the BACTEC Mycobacteria Growth Indicator Tube 960 (MGIT) and Ogawa media among patients with tuberculous pleurisy. Out of 104 effusion samples, 43 (41.3%) were culture positive on either the MGIT or the Ogawa media. The culture yield of MGIT was higher (40.4%, 42/104) than that of Ogawa media (18.3%, 19/104) (P<0.001). One of the samples was positive only on the Ogawa medium. The median time to positivity was faster in the MGIT (18 days, range 8-32 days) than in the Ogawa media (37 days, range 20-59 days) (P<0.001). No contamination or growth of nontuberculous mycobacterium was observed on either of the culture media. In conclusion, the automated liquid culture system could provide approximately twice as high yields and fast results in effusion culture, compared to solid media. Supplemental solid media may have a limited impact on maximizing sensitivity in effusion culture; however, further studies are required.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Automation, Laboratory/*methods ; Cell Culture Techniques ; Culture Media/*classification ; Female ; Humans ; Male ; Middle Aged ; Mycobacterium tuberculosis ; Pleura/microbiology/pathology ; Retrospective Studies ; Sputum/*microbiology ; Tuberculosis, Pleural/*diagnosis ; Young Adult

No abstract available.
Automation, Laboratory/*methods ; Culture Media/*classification ; Female ; Humans ; Male ; Sputum/*microbiology ; Tuberculosis, Pleural/*diagnosis

Succinic acid is an important C4 platform chemical in the synthesis of many commodity and special chemicals. In the present work, different compounds were evaluated for succinic acid production by Actinobacillus succinogenes GXAS 137. Important parameters were screened by the single factor experiment and Plackeet-Burman design. Subsequently, the highest production of succinic acid was approached by the path of steepest ascent. Then, the optimum values of the parameters were obtained by Box-Behnken design. The results show that the important parameters were glucose, yeast extract and MgCO3 concentrations. The optimum condition was as follows (g/L): glucose 70.00, yeast extract 9.20 and MgCO3 58.10. Succinic acid yield reached 47.64 g/L at the optimal condition. Succinic acid increased by 29.14% than that before the optimization (36.89 g/L). Response surface methodology was proven to be a powerful tool to optimize succinic acid production.
Actinobacillus ; classification ; genetics ; metabolism ; Bioreactors ; Culture Media ; metabolism ; Fermentation ; Glucose ; metabolism ; Industrial Microbiology ; methods ; Succinic Acid ; metabolism

In order to lower the cost of lipid production of microalgae and reduce greenhouse gas emissions, microalgae Chlorococcum alkaliphilus MC-1 with the characteristics of rapid pH drift and high pH adaptability, was cultivated with bubbling of flue gas. The experiment was first performed in the photobioreactor (15 L) in three groups (control group, CO2 group and flue gas group), then, in the open raceway pond (24 m2). The adaptability of microalgae MC-1 to the cultivation with flue gas was studied. The results showed that the maximum biomass concentration, growth rate, total lipid content and CO2 fixation rate were (1.02+/-0.07) g/L, (0.12+/-0.02) g/(L.d), (37.84+/-0.58)% and (0.20+/-0.02) g/(L.d) in the photobioreactor treated with flue gas, 36%, 33.33%, 15.34% and 33.33% higher than those of the CO2 group, respectively. In the open raceway pond with aeration of flue gas, the maximum biomass concentration, growth rate, total lipid content and CO2 fixation rate were 147.40 g/m2, 14.73 g/(m2.d), 35.72% and 24.01 g/(m2.d), respectively, which were similar to the cultivation with pure CO2. The toxic heavy metal contents (Pb, As, Cd and Cr) in the biomass of MC-1 treated with flue gas were all below the legal limits. Additionally, the absorptive effect of CO2, NO and SO2 were determined. In the photobioreactor and open raceway pond, the average absorption ratios of these gases were all higher than previous studies. Therefore, our study showed that MC-1 can adapt to the cultivation with flue gas, and it is feasible to enlarge the outdoor cultivation of MC-1 for lipid production coupling with emissions reduction of flue gas.
Adaptation, Physiological ; physiology ; Carbon Dioxide ; chemistry ; Chlorophyta ; classification ; growth & development ; physiology ; Culture Media ; metabolism ; Culture Techniques ; methods ; Gases ; chemistry ; Microalgae ; classification ; growth & development ; physiology ; Nitric Oxide ; chemistry ; Sulfur Dioxide ; chemistry

OBJECTIVEThe research aimed to investigate the entophytic fungal community of Cynanchum Komarrovii, including the biodiversity in different organs and the correlations with ecological environment. Endophytic fungi with patent bioactivity were also rapidly screened.

METHODPDA medium was used to isolate and purify the endophytic fungi from C. komarovii living in Shaanxi and Ningxia district, respectively. The strains were identified based on the morphological characteristics of the fungi and similarity of 5.8S gene and internal transcribed spacer (ITS) sequence. Pyriculaia oryzae model was applied to preliminarily screen the active fungi.

RESULTNinety-four strains of endophytic fungi were isolated and identified to 9 species, 13 genera, 9 families and 6 orders, among them, 47 strains were from the plants living in Ningxia. And then, 5 of them were isolated from roots, 14 from branches, and 28 from leaves. They were identified belonging to 8 species, 9 genera, 5 families and 4 orders. Additionally, 47 strains were from the plants living in Shaanxi. 16 were isolated from the roots, 18 from branches, 13 from leaves. They were identified belonging to 5 species, 8 genera, 6 families and 4 orders. By preliminary screening, 18 strains of endophytes completely inhibited the germination of conidium, which showed a potential bioactivity for these fungi. Both N4 and S17 strains had stronger growth inhibition effect.

CONCLUSIONEndophytic fungi from desert plant C. komarovii have the feature of diversity. Different geographical environment and type of organizations lead to the significant difference on the quantity and the species composition. Most of fungi in Ningxia C. komarovii distribute in leaves. However, most of those in Shaanxi C. komarovii distribute in stems and leaves. It also indicated that endophytes from C. komarovii had a strong antifungal activity.

Antifungal Agents ; pharmacology ; Biodiversity ; China ; Culture Media, Conditioned ; pharmacology ; Cynanchum ; microbiology ; DNA, Ribosomal Spacer ; genetics ; Desert Climate ; Endophytes ; classification ; genetics ; isolation & purification ; Fungi ; classification ; genetics ; isolation & purification ; Genetic Variation ; Magnaporthe ; drug effects ; growth & development ; Microbial Sensitivity Tests ; Phylogeny ; Plant Leaves ; microbiology ; Plant Roots ; microbiology ; Plant Stems ; microbiology ; RNA, Ribosomal, 5.8S ; genetics ; Species Specificity

Five Klebsiella pneumonia strains (including two strains whose genes for lactic acid were knocked out) were used to produce 2,3-butanediol, in which K. pneumonia HR521 LDH (gene for lactic acid was knocked out) was the best for the production, and then the fermentation medium was optimized by orthogonal design. The optimum compositions were as follows: glucose 90 g/L, (NH4)2HPO4 3 g/L, CLSP 6 g/L, sodium acetate 5 g/L, KCl 0.4 g/L, MgSO4 0.1 g/L, FeSO4 x 7H2O 0.02 g/L, MnSO4 0.01 g/L. Under the above conditions, final concentration of acetone and 2,3-butanediol could reach 37.46 g/L, 10 g/L higher than that under the initial conditions, the yield was 90.53% of the theory, and the productivity was 1.5 g/(L-h), and no lactic acid was detected, which could be benefit for the downstream processing and industrial application.
Butylene Glycols ; metabolism ; Culture Media ; chemistry ; Fermentation ; Gene Knockout Techniques ; Glucose ; metabolism ; Klebsiella pneumoniae ; classification ; genetics ; growth & development ; metabolism

To investigate the energy utilization efficiency of Synechococcus sp. PCC7942 under mixotrophic conditions, we studied its growth characteristics in mixotrophic cultures with glucose and acetic acid respectively and discussed the carbon metabolism and energy utilization based on metabolic flux analysis. Results showed that both glucose and acetate could better enhance the growth of Synechococcus sp. PCC7942, and the latter was more effective. The metabolic flux through glycolytic pathway in mixotrophic cultures was stimulated by glucose whereas depressed by acetate, while the flux through the tricarboxylic acid cycle increased in both cases. Under mixotrophic conditions, glucose makes more significant impact on the diminishment of photochemical efficiency of Synechococcus sp. PCC7942. Although the contribution of light energy was smaller, the cell yields based on total energy in mixotrophic cultures were higher comparing with photoautotrophic culture. The energy conversion efficiencies based on ATP synthesis in photoautotrophic culture, mixotrophic cultures with glucose and with acetate were evaluated to be 6.81%, 7.43% and 8.77%, respectively.
Acetic Acid ; pharmacology ; Adenosine Triphosphate ; biosynthesis ; Carbon ; metabolism ; Culture Media ; Culture Techniques ; methods ; Energy Metabolism ; Glucose ; pharmacology ; Synechococcus ; classification ; growth & development ; metabolism

BACKGROUND: Procedures for rapid identification and susceptibility testing by direct inoculation (DI) from positive blood culture bottles into an automated system have not been standardized. This study was purposed to evaluate DI from BACTEC 9240 blood culture system (BD, USA) into MicroScan (Dade Behring, USA) or Phoenix (BD, USA). METHODS: From May to June 2006, bacterial pellets from positive aerobic bottles showing gram-positive cocci (GPC) or gram-negative rods (GNR) of single morphology were directly inoculated to MicroScan PosCombo1A and NegCombo32 and to Phoenix PMIC/ID-107 and NMIC/ID-53. In addition, the automated instruments were also inoculated from subcultures (standard inoculations, SI). Species identification and susceptibilities were compared between DI and SI and between MicroScan and Phoenix. RESULTS: A total of 108, 104, and 78 specimens were tested with MicroScan, Phoenix, and both, respectively. When DI and SI were matched, 94.8% of GPC were correctly identified with MicroScan, compared to 80.7% with Phoenix, and 93.9% of GNR were correctly identified with MicroScan, compared to 95.7% with Phoenix. DI with MicroScan and Phoenix showed correct susceptibilities in 94.6% of 1,150 and 96.5% of 660 tests (with very major error [VME] of 1.1% and 1.1%), respectively, among GPC and in 94.4% of 942 and 96.3% of 781 tests (with VME of 0.6% and 0%), respectively, of GNR. Correlation of identification/susceptibilities between MicroScan and Phoenix using DI were 81.8%/98.0% for Staphylococcus aureus and 100.0%/95.6% for Escherichia coli. CONCLUSIONS: DI warrants a reliable method for identification and susceptibility testing of both GPC and GNR in MicroScan, and those of only GNR in Phoenix.
Automation ; Bacterial Typing Techniques/instrumentation/*methods ; Culture Media ; Gram-Negative Bacteria/*classification/drug effects/isolation & purification ; Gram-Negative Bacterial Infections/blood/*microbiology ; Gram-Positive Bacterial Infections/blood/*microbiology ; Gram-Positive Cocci/*classification/drug effects/isolation & purification ; Humans ; Microbial Sensitivity Tests/instrumentation/*methods ; Reagent Kits, Diagnostic ; Sensitivity and Specificity

A beta-D-xylosidase from Leifsonia shinshuensis DICP 16 was purified to apparent homogeneity using a combination of ammonium sulfate precipitation, DE 52 anion-exchange, Q-Sepharose Fast Flow anion-exchange, Toyopearl Butyl 650C hydrophobic-interaction and Sephacryl S-300 HR gel-permeation chromatography. The purified xylosidase consisted of two same subunits and had the relative molecular weight of 180 kD as determined by SDS-PAGE and gel-permeation chromatography. The maximal beta-D-xylosidase activity occurred at 55 degrees C and pH 7.0. It was stable at 45 degrees C and retained its original activity for 60 min. The stability declined rapidly when the temperature rose above 55 degrees C. The xylosidase was stable in the pH range from 6.0 to 11.0 for 20 h. At pH 7.0 and 45 degrees C the Km for p-nitrophenyl-beta-D-xylopyranoside (pNPX) was 1.04 mmol/L and the Vmx was 0.095 mmol nitrophenol/min/mg xylosidase. The enzyme was inhibited strongly by Fe2+ and Cu2+. It exhibited low levels of activity against other artificial substrates, compared to its activity against pNPX. When different natural xylosides were used as the substrates, the xylosidase showed distinct hydrolysis ability. It could hydrolyze 20-C, beta-(1-->6)-xyloside of ginsenoside Rb3 (G-Rb3) into ginsenoside Rd, but did not hydrolyze the other beta-D-glucosidic bonds of G-Rb3. Additionally, the xylosidase could not hydrolyze C-7 xylosyl-bearing taxanes.
Actinomycetales ; classification ; enzymology ; Amino Acid Sequence ; Culture Media ; Hydrogen-Ion Concentration ; Molecular Sequence Data ; Sequence Analysis, Protein ; Temperature ; Xylosidases ; chemistry ; genetics ; isolation & purification